Combination optical hemoglobin and electrochemical lead assay

ABSTRACT

A sensor and analyzer for measuring an analyte in a liquid sample are disclosed. The sensor includes a substrate with a reservoir disposed therein. The reservoir may include a top surface and a bottom surface, at least one transparent portion forming at least a part of the bottom surface of the reservoir, and a reflector disposed on the upper surface of the reservoir at a location opposite the at least one transparent portion. The analyzer may include a support surface, an aperture extending through the support surface, a light source disposed below the support surface and oriented so that at least a portion of the light emitted from the light source passes through the aperture, and a detector configured to measure an intensity of light received at the detector.

INCORPORATION BY REFERENCE TO ANY PRIORITY APPLICATIONS

This application claims is a continuation of U.S. application Ser. No.14/978,292, filed Dec. 22, 2015, which claims priority to U.S.Provisional Application No. 62/096,178, filed Dec. 23, 2014, both ofwhich are incorporated herein by reference. Any and all applications forwhich a foreign or domestic priority claim is identified in theApplication Data Sheet as filed with the present application are herebyincorporated by reference under 37 CFR 1.57.

BACKGROUND Field

This disclosure relates to a sensor, analyzer, and method for analyzingat least one analyte in sample.

Description

It is frequently desired to analyze the amount of an analyte in a liquidsample, such as blood or other biological fluids. When sampling analytesin blood, it may be desirable to sample for more than one analyte. Thismay require using a separate sensor, reagent, and/or sampling apparatusfor each analyte. However, using a separate sensor, reagent, and/orsampling apparatus may be time consuming and costly. Therefore, a sensorwhich can detect and/or measure the concentration of two analytes in asingle sampling operation may be desired.

Electrochemical stripping and square wave voltammetry techniques havebeen developed using colloidal gold based sensors to measureconcentrations of various analytes, such as lead, in a blood sample.Some exemplary techniques and apparatuses are described in U.S. Pat. No.5,873,990, entitled “Handheld Electromonitor Device,” the entirety ofwhich is incorporated herein by reference. These techniques allow forlow cost, quick, and accurate testing of blood lead concentration; theydo not, however, test for any additional analytes.

A correlation between the color of blood samples treated withhydrochloric acid and hemoglobin concentration has long been observed.The Sahli hemoglobin method, developed in the early 1900s and still usedtoday in some parts of the world, estimates blood hemoglobinconcentration by matching the color of treated blood to predeterminedsamples and/or color standards. This method however, remains impreciseand fails to test for any additional analytes.

SUMMARY

In one aspect, a sensor for measuring an analyte in a liquid sample isdisclosed. The sensor includes a reservoir having a top surface and abottom surface, at least one transparent portion forming at least a partof the bottom surface of the reservoir, and a portion of the top surfacethat comprises a reflector.

In some embodiments, the substrate further comprises a base layerforming the bottom surface of the reservoir, wherein the at least onetransparent portion forms at least a portion of the base layer; a firstspacer layer having a first void extending through a thickness of thefirst spacer layer; a second spacer layer having a second void extendingthrough a thickness of the second spacer layer and wherein at least aportion of a bottom surface of the second spacer layer comprises theportion of the top surface of the reservoir comprising the reflector; alid having a bottom surface, and wherein at least a portion of thebottom surface of the lid forms at least a portion of the top surface ofthe reservoir. The first spacer layer is disposed on the base layer, thesecond spacer layer is disposed on the first spacer layer, and the lidis disposed on the second spacer layer. In some embodiments, thereservoir further comprises a first depth between the bottom surface ofthe reservoir and a first portion of the upper surface of the reservoir,and a second depth between the bottom surface of the reservoir and asecond portion of the upper surface of the reservoir, wherein the firstdepth is less than the second depth, and wherein the reflector isdisposed on the first portion of the upper surface of the reservoir. Insome embodiments, the first depth is equal to the thickness of the firstspacer layer, and the second depth is equal to a combined thickness ofthe thickness of the first spacer layer and the thickness of the secondspacer layer. In some embodiments, the first depth and the second depthmay also include a thickness of one or more adhesive layers. In someembodiments, the sensor is configured to be used to analyze for ahemoglobin concentration of the liquid sample using an opticalmeasurement.

In some embodiments, the sensor may further comprise at least oneelectrode disposed on a bottom surface of the reservoir and at least oneelectrical contact disposed on the base layer, wherein the at least oneelectrode is in electrical communication with the at least oneelectrical contact. In some embodiments, a first of the at least oneelectrodes comprises a colloidal gold deposit. In some embodiments, thesensor is configured to be used to analyze for a hemoglobinconcentration of the liquid sample using an optical measurement and toanalyze for lead concentration using an electrochemical measurement. Insome embodiments, the liquid sample is a blood sample treated withhydrochloric acid.

In a second aspect, an analyzer for measuring an analyte in a liquidsample is disclosed. The analyzer comprises a port for receiving asensor and having a support surface configured to support the sensor, anaperture extending through the support surface, a light source disposedbelow the support surface and oriented so that at least a portion of thelight emitted from the light source passes through the aperture, adetector configured to measure an intensity of light received at thedetector; and a processor electrically coupled to the detector toreceive an output of the detector.

In some embodiments, the analyzer further comprises a window disposedwithin the aperture. In some embodiments, the window comprises sapphire.In some embodiments, the detector is disposed below the support surfaceof the analyzer. In some embodiments, the light source comprises firstand second light sources, and the first and second light sources areconfigured to alternatingly emit light. In some embodiments, the firstand second light sources further comprise integrated lenses configuredto focus the light emitted through the aperture. In some embodiments,the first and second light sources are configured so that the lightemitted from each passes through the aperture at an approximately 45°angle relative to a central axis of the aperture. In some embodiments,the light source and detector are configured to emit and detect light ata wavelength corresponding to an isosbestic point of the liquid sample.In some embodiments, the light source and detector are configured toemit and detect light with a wavelength of approximately 405 nm whichrepresents an isosbestic point of a blood sample treated withhydrochloric acid. In some embodiments, the analyzer further comprises aclock, the clock electrically connected to the light source and thedetector, and configured so that the light source can be pulsed at afirst frequency and the detector can be demodulated at the firstfrequency.

In some embodiments, the analyzer is configured to make a first opticalmeasurement of light reflected off a reflector of the sensor before theliquid sample is introduced and a second optical measurement of lightreflected off a reflector of the sensor after the liquid sample isintroduced.

In a third aspect, a method for measuring an analyte in a liquid sampleis disclosed. The method comprises inserting a sensor into an analyzer;introducing the liquid sample to a reservoir in the sensor, illuminatingthe liquid sample in the sensor using a light source in the analyzer,measuring a reflectance of the liquid sample using a detector in theanalyzer, and computing a measurement of the analyte using the measuredreflectance.

In some embodiments, the reflectance is measured by measuring lightreflected off a reflective surface in the sensor. In some embodiments,the reflectance is computed by comparing an intensity measured at thedetector to a reference intensity. In some embodiments, the referenceintensity is obtained by inserting an empty sensor into the analyzer,illuminating the empty sensor, and measuring an intensity of lightreceived at the detector. In some embodiments, internally reflectedstray light is measured by detecting at the detector the intensity oflight reflected off a light absorbing surface as the sensor is insertedinto or withdrawn from the analyzer, and the method further comprisessubtracting the measured internally reflected stray light from thereference intensity and the measured intensity of the sample to obtain aresult which corrects for internally reflected stray light.

In some embodiments, the measured analyte is hemoglobin and the liquidsample comprises a blood sample treated with hydrochloric acid. In someembodiments, the method further comprises making an electrochemicalmeasurement of lead using the same sensor and analyzer.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 depicts a perspective view of an embodiment of a combinationelectrochemical lead and optical hemoglobin sensor.

FIG. 2 depicts each of the layers individually of an embodiment of acombination electrochemical lead and optical hemoglobin sensor.

FIGS. 3A and 3B depict embodiments of electrical contacts disposed on abase layer of a sensor.

FIG. 4 depicts an embodiment of a second spacer layer including only asingle reservoir space.

FIGS. 5A and 5B depict simplified longitudinal cross-section views ofembodiments of a combination electrochemical lead and optical hemoglobinsensor taken along the line A-A′ in FIGS. 2 and 4.

FIG. 6 depicts a simplified longitudinal cross-section view of anembodiment of a combination electrochemical lead and optical hemoglobinsensor.

FIG. 7 depicts a perspective view of an embodiment of an analyzer with acombination electrochemical lead and optical hemoglobin sensor.

FIG. 8 depicts a perspective view of an embodiment of a sensor supportstructure and an optical system housing.

FIG. 9 depicts a cross-sectional view of an embodiment of an opticalanalyzer.

FIG. 10 depicts a perspective view of an embodiment of components of anoptical system with the optical system housing removed.

FIG. 11 depicts a simplified view of an embodiment of the operation ofan optical system capable of lock-in detection.

FIG. 12 is a graph showing a linear relationship between opticalabsorbance and hemoglobin concentration at three light wavelengths.

FIG. 13 depicts a simplified view of the operation of an embodiment ofan optical system.

FIG. 14 depicts exemplary optical measurements of a sensor in bothfilled and empty states.

FIG. 15 depicts optical measurements taken with no sensor in place, withan empty sensor in place, and with a filled sensor in place.

FIG. 16 is a graph of an example of a measured reflectance signal takenas a sensor is inserted into the analyzer.

FIG. 17 is a graph depicting two non-linear curves relating measuredreflectance to hemoglobin concentration.

FIG. 18 is a graph which indicates how the absorbance of a treated bloodsample may change over time when measured optically at differentwavelengths of light.

FIG. 19 is a graph comparing the results of optical hemoglobinmeasurements taken using the principles of the present disclosure withreference measurements.

DETAILED DESCRIPTION

In the following detailed description, reference is made to theaccompanying drawings, which form a part hereof. In the drawings,similar symbols typically identify similar components, unless contextdictates otherwise. The illustrative embodiments described in thedetailed description, drawings, and claims are not meant to be limiting.Other embodiments may be utilized, and other changes may be made,without departing from the spirit or scope of the subject matterpresented herein. It will be readily understood that the aspects of thepresent disclosure, as generally described herein, and illustrated inthe Figures, can be arranged, substituted, combined, separated, anddesigned in a wide variety of different configurations, all of which areintended to be within the scope of this disclosure.

Disclosed in the present application are a sensor, analyzer, system, andmethods for analyzing a sample for at least one analyte. In someembodiments, the sample is a vertebrate or mammalian blood sample, andthe sample is placed on the sensor of the present disclosure, the sensorbeing readable using the analyzer. In some embodiments, the sample maybe treated with a reagent to facilitate the analysis. In someembodiments, the reagent is hydrochloric acid. In some embodiments, thesample is analyzed for hemoglobin concentration and results may beprovided to a user in grams of hemoglobin per deciliter of sample(g/dL). In some embodiments, the sample is analyzed for leadconcentration and hemoglobin concentration. A single sensor may be usedto analyze the sample for lead concentration using an electrochemicalmeasurement and may further be used to analyze the sample for hemoglobinconcentration using an optical measurement. A sensor having a substratesuitable for use in sampling blood lead levels is described in U.S. Pat.No. 5,468,366, entitled “Colloidal-Gold Electrosensor Measuring Device,”the entire contents of which are herein incorporated by reference. Bloodlead concentration analysis can be performed using systems and methodssimilar to those described in U.S. Pat. No. 5,873,990, referenced above.

As used herein, the terms “simultaneously” or “at the same time” neednot necessarily mean at exactly the same moment, and may mean that twoactions or operations occur concurrently. For example, in the followingdisclosure reference is made to analyzing lead concentration andhemoglobin concentration at the same time or simultaneously. This neednot mean that the sensors or sampling apparatus are performing theanalysis at exactly the same moment, or that electrical or opticalsignals are applied to the sensing electrodes or detectors of the sensorat exactly the same instant. Simultaneous measurement or measurement “atthe same time” of lead and hemoglobin may mean that lead and hemoglobinare measured using a single sensor and sampling apparatus, or by testingthe same blood sample, possibly with little to no need for operatorintervention between the analyses. The measurement of lead andhemoglobin may occur sequentially, such as lead measurement first, andthen hemoglobin measurement, or vice versa, at generally the same time,or within a short time window. In some embodiments, the sensor may beused to make only a single measurement of one analyte.

FIG. 1 depicts an embodiment of a combination electrochemical lead andoptical hemoglobin sensor configured to receive a liquid sample andfacilitate analysis of at least one analyte in the sample. The sensor100 is generally rectangular in shape and may comprise base layer 110and a lid layer 140 disposed on the base layer. Lid layer 140 includes asample inlet 141 and a vent 142, each formed as holes that extendthrough a thickness of lid layer 140. In some embodiments, other layersmay be disposed between base layer 110 and lid layer 140. Sensor 100 mayfurther comprise an overall length dimension measured between the firstend 101 and second end 102 along a line perpendicular to first end 101;an overall width dimension, measured along first end 101 or second end102; and an overall thickness dimension, measured between a top surfaceof lid layer 140 and a bottom surface of base layer 110 along a linenormal to a top surface of lid layer 140. In some embodiments, theoverall length dimension is about 1.72″ the overall width dimension isabout 0.55″, and the overall thickness dimension is about 0.031″. Itwill be understood by one of skill in the art, according to theprinciples and embodiments presently disclosed, that other dimensionsare possible and within the scope of the present disclosure. Forexample, in some embodiments the overall length dimension is betweenabout 0.5″ and about 6″, the overall width dimension is between about0.25″ and about 3″, and the overall thickness is between about 0.005″and about 0.5″; however, other sizes outside of these ranges arepossible and contemplated. Further, it should be noted that othershapes, besides rectangular, may be used according to the principles andsubject matter presently disclosed. For example, in some embodiments,sensor 100 may be substantially circular. In some embodiments, thedimensions of the sensor may correspond to a sensor port on an analyzerwhich will be described in greater detail below.

In some embodiments, sensor 100 comprises first end 101 and second end102. First end 101 includes a plurality of contacts 111-114 and isconfigured in size and shape to be insertable into a sample port on ananalyzer, wherein the sample port has a compatible geometry configuredto receive first end 101 of sensor 100. In some embodiments, the crosssection of the sensor 100 and the sample port are substantiallyrectangular. Contacts 111-114 will be discussed in greater detail below.In some embodiments, sensor 100 is configured so that second end 102remains exposed when first end 101 has been inserted into the analyzer.This may allow a user to introduce the liquid sample to sensor 100 aftersensor 100 has been inserted into the analyzer.

Referring now to FIG. 2, sensor 100 includes several layers stacked ontop of each other to form the various features of sensor 100. Sensor 100may comprise a base layer 110, a first spacer layer 120, a second spacerlayer 130, and a lid layer 140. A thin layer of adhesive may be appliedbetween each successively stacked layer, bonding the layers together toform sensor 100. In some embodiments, each layer of adhesive isapproximately 0.001 inches thick, although it will be understood by oneof skill in the art that different thicknesses may be used. In someembodiments, bonding methods other than adhesive may be used, or sensor100 may be manufactured or formed as a unitary piece, either throughprinting, molding, or other suitable manufacturing process.

Each layer of sensor 100 will now be described in greater detail withreference to FIG. 2, which depicts embodiments of each of the layersindividually for convenience and ease of description. In someembodiments, base layer 110 is generally rectangular in shape having alength of approximately 1.72″ a width of approximately 0.55″, and athickness of approximately 0.01″; it will be understood by one of skillin the art, however, that other dimensions for the base layer may beused. In some embodiments the length of base 110 extends beyond theother layers in a longitudinal direction, each end of base layer 110forming one of first end 101 and second end 102 of sensor 100.

Base layer 110 may comprise a transparent substrate that permits opticalsignals to pass there through. In some embodiments, the base layer 110is formed entirely of a transparent material. In some embodiments, baselayer 110 is only partially comprised of a transparent material, thetransparent material forming a transmission window 119 through the baselayer 110 to allow for optical interrogation of a sample. In someembodiments, the transmission window 119 is disposed between a workingelectrode 116 and a counter electrode 117 along a longitudinal axis ofbase layer 110. The optically transparent material of base layer 110 maybe formed from plastic, glass, or other suitable material that permitslight of the wavelengths discussed below to be transmitted therethrough. In some embodiments, at least the transmission window 119 ofthe base layer 110 is made from polycarbonate or polyester. In someembodiments of base layer 110 a hard-coated, optical grade polycarbonatewith a gloss finish is used for the transmission window 119.

The components used to make an electrochemical measurement of the leadconcentration of a liquid sample are disposed on an upper surface ofbase layer 110. These include contacts 111-114, traces 111 a-113 a, andelectrodes 115-117. In some embodiments, the contacts 111-114 and traces111 a-113 a are a silver-containing material screen printed onto baselayer 110. In some embodiments, contacts 111-114 and traces 111 a-113 ainclude a carbon layer screen printed on top of the silver layer. Insome embodiments, the contacts and electrical traces may be printed,etched, or otherwise deposited on the base layer 110.

In the illustrated embodiment, sensor 100 includes four contacts: aworking electrode contact 111, an auxiliary or counter electrode contact112, a reference electrode contact 113, and a sensor insertion contact114. The contacts 111-114 are disposed on first end 101 of sensor 100 onan upper surface of base layer 110, and are exposed such that uponinsertion of sensor 100 into a sample port of an analyzer, the contacts111-114 make physical contact with corresponding contacts in theanalyzer forming an electrical connection between sensor 100 and theanalyzer.

Each of contacts 111-113 is in electrical communication with traces 111a-113 a, respectively, the traces 111 a-113 a extending generally awayfrom the first end 101 of sensor 100 and toward the second end 102 ofsensor 100. Through trace 111 a, the working electrode contact 111 is inelectrical contact with the working electrode 116. Through trace 112 a,the counter electrode contact 112 is in electrical contact with thecounter electrode 117. Through trace 113 a, reference electrode contact113 is in electrical contact with the reference electrode 115. Althoughone configuration is depicted in FIG. 2 for the electric traces and thecontacts, one of skill in the art will understand that a differentcontact order or trace configuration can be used without departing fromthe scope of the present application (see, for example, FIGS. 3A and3B).

Working electrode 116 is disposed on an upper surface of base 110 andincludes a layer of carbon which has been sputtered, printed, sprayed,air brushed, or otherwise deposited on base layer 110. The workingelectrode may also advantageously comprise a colloidal gold solutionsputtered, printed, sprayed, air brushed, or otherwise deposited on thecarbon layer. The counter electrode 117 is similarly disposed on baselayer 110. Counter electrode 117 may be comprised of carbon and may beformed through the same processes described in reference to the workingelectrode 116. Reference electrode 115 may comprise carbon, silver, orsilver chloride and is similarly disposed on base layer 110. Electrodes115-117 are disposed on an upper surface of base layer 110 so as to comeinto contact with a sample in the sensor 100. It will be appreciated byone of skill in the art that the particular arrangement, order, materialof construction, and/or number of the electrodes 115-117 may varywithout departing from the scope of the present disclosure. The functionof each electrode 115-117 is discussed elsewhere.

The first spacer layer 120 is disposed on an upper surface of base layer110. First spacer layer 120 may also be generally rectangular in shapewith a width less than or equal to the width of the base layer 110 and alength less than the length of base layer 110. First spacer layer 120may advantageously be about 0.002″ thick, and include a 0.001″ thicklayer of adhesive on each side for a total thickness of about 0.004″;although one of skill in the art will understand according to thepresent disclosure that other thicknesses may be used, for example,thicknesses of approximately 0.0005″, 0.001″, 0.005″, 0.010″, or anythickness there between. As will be described in greater detail belowwith regard to FIGS. 5A-5B, the thickness of the first spacer layer 120affects the path length of light traveling through the sample andaffects the amount of light available for detection.

First spacer layer 120 is disposed on top of base layer 110 so that thecontacts 111-114, also disposed on base layer 110, remain exposed. Asseen in FIG. 2, first spacer layer 120 includes a first sample reservoirspace 123 that is formed as a void in first spacer layer 120. The firstsample reservoir space 123 is configured in size and shape to surroundthe electrodes 115-117 when both the first spacer layer 120 and theelectrodes 115-117 are disposed on base layer 110. First spacer layer120 may also include an inlet portion 121 configured in size and shapeto align with sample inlet 141 when sensor 100 is fully assembled. Thefirst spacer layer 120 may comprise an electrically insulating material,such as Mylar®, or other similar material. In some embodiments, thematerial may be hydrophilic, or coated with a hydrophilic substance.

The second spacer layer 130 is disposed on top of the first spacer layer120. Second spacer layer 130 may have the same width and lengthdimensions as the first spacer layer 120. Second spacer layer 130 may bemade from white polyester or any other suitable material. In someembodiments, a suitable material may be one that can be used as adiffuse reflector. In some embodiments, the material may be hydrophilic,or coated with a hydrophilic substance. The combined thickness of firstspacer layer 120 and second spacer layer 130 defines a depth of areservoir within sensor 100 that allows electrodes 115-117 to be used tomake an electrochemical measurement of lead concentration. This depthwill be discussed in greater detail below with reference to FIGS. 5A-5B.In some embodiments the second spacer layer 130 is approximately 0.001,0.005, 0.01, 0.15, 0.2 inches thick or more, or any thicknesstherebetween. The thickness of the second spacer layer 130 affects thevolume of sample, such as blood, which is accommodated on the sensor. Aperson of skill in the art, guided by the present disclosure, willunderstand how to vary the thickness of the first spacer layer 120 andthe second spacer layer 130 in order to obtain an electrochemical leadmeasurement at the working electrode 115.

In some embodiments, second spacer layer 130 includes a second samplereservoir space 133 and a third sample reservoir space 135, each formedas voids in second spacer 130. Second and third sample reservoir spaces133, 135 are separated by a bridge 136. The bridge 136 includes theportion of the second spacer layer 130 located between the second andthird reservoirs 133 and 135, and may be formed as an integral piece ofthe second spacer layer 130.

In some embodiments, the bridge 136 itself may comprise the reflector137; for example, when second spacer layer 130 including bridge 136 ismade from white polyester, which itself acts as a diffuse reflector, noadditional reflector is needed. When second spacer layer 130 isassembled on top of first spacer layer 120, the second sample reservoirspace 133 and the third sample reservoir space 135 are in fluidcommunication with each other by means of the first sample reservoirspace 123 of the first spacer layer 120, as shown in FIG. 5A. Secondspacer layer 130 may also include an inlet portion 131 configured insize and shape to align with sample inlet 141 when sensor 100 is fullyassembled.

A lid layer 140 is disposed on top of second spacer layer 130. The lidlayer 140 is configured in size and shape to have the same width andlength dimensions as second spacer layer 130. In some embodiments, lidlayer 140 is about 0.001, 0.005, 0.01, 0.02 inches thick or more, or anyvalue there between. Lid layer 140 may be comprised of a plastic orother suitable material. In some embodiments, lid layer 140 is coatedwith a hydrophilic substance so that the reservoir can be more easilyfilled with the sample. In some embodiments, lid layer 140 may also beformed of a clear, transparent, or translucent material which provides avisual indication to the user when the reservoir is filled. In someembodiments, lid layer 140 may be opaque so as to shield the opticalmeasurements that will be discussed below from interference from ambientlight. It will be noted, however, that a clear lid layer 140 may be usedand obtain an accurate optical measurement according to the presentdisclosure. The lid layer 140 provides an upper boundary on a samplereservoir within sensor 100 to prevent evaporation of the sample. Lidlayer 140 also includes a sample inlet 141 and a vent 142 formed asvoids extending through a thickness of the lid layer 140. The relativepositioning of the inlet 141 and vent 142 depicted in FIG. 2 is merelyillustrative and one of skill in the art will appreciate that thepositioning of the inlet 141 and vent 142 may vary without departingfrom the scope of the present disclosure.

FIGS. 3A and 3B each depict a sensor 100 with an embodiment of a layoutof five contacts: a working electrode contact 111, an auxiliary orcounter electrode contact 112, a reference electrode contact 113, asensor insertion contract 114, and a sensor identifier contact 114a. Asshown, some of the contacts disposed on first end 101 may be spaced backfrom the edge of first end 101, for example, contacts 111, 112, 113.Other contacts may be disposed directly on the edge, for example,contacts 114, 114a. Moreover, in some embodiments, the lengths andwidths of the contracts may vary from contact to contact. In someembodiments, greater than five or fewer than four contacts may be used.

FIG. 4 depicts an embodiment of the second spacer layer 130 which doesnot include third sample reservoir space 135. Third sample reservoirspace 135 is omitted and bridge 136 and reflector 137 have beenenlarged. This will be described in greater detail below. In someembodiments, second spacer layer 130 may include a vent 132.

FIGS. 5A and 5B depict simplified (not to scale) longitudinalcross-sectioned views of embodiments of an assembled sensor 100 takenalong the lines A-A′ shown on the individual layers in FIGS. 2 and 4. Asshown in FIG. 5A, the various layers of sensor 100 define an internalsample reservoir 151. The sample reservoir 151 includes the first samplereservoir space 123 (shown in dashed lines) defined at its lateral edgesby first spacer 120, the second sample reservoir space 133 (shown indashed lines), and the third sample reservoir space 135 (shown in dashedlines). The second sample reservoir space 133 and the third samplereservoir space 135 are both defined at their lateral edges by thesecond spacer layer 130. The thickness of first spacer layer 120 alongwith the thickness of the adhesive that binds this layer to the adjacentlayers define the depth of the first reservoir space 123, which impactsthe sensor's ability to be used for optical hemoglobin measurement. Thethickness of the second spacer layer 130 defines the depth of the secondsample reservoir space 133 and the third sample reservoir space 135.

The sample to be analyzed is introduced to sensor 100 at sample inlet141, filling sample reservoir 151, including the first sample reservoirspace 123, the second sample reservoir space 133, and the third samplereservoir space 135. Vent 142 is provided to prevent overfilling and toallow air to escape as sample reservoir 151 is filled.

In some embodiments, for example as shown in FIG. 5B, the third samplereservoir space 135 is omitted from second spacer layer 120, as shown inFIG. 4. The third sample reservoir space 135 may be omitted by extendingbridge 136. Omitting the third reservoir space 135 may improve fillingof the reservoir. Accordingly, sample reservoir 151 may comprise onlyfirst sample reservoir space 123 and second sample reservoir 133. A vent132, 142 may also be included.

As shown in FIGS. 5A and 5B electrodes 115-117 are disposed on baselayer 110 and are oriented so as to come into contact with a sample insample reservoir 151. The bottom surface of bridge 136 of second spacer130 may serve as a diffuse reflector 137, which is used in determiningthe hemoglobin concentration of the sample and will be more fullydescribed below. Reflector 137 is disposed on bridge 136 of secondspacer 130 at a location so as to be in contact with the sample insample reservoir 151. Further, transmission window 119 is disposed onbase layer 110 at a location substantially opposite the reflector 137.The transmission window 119 should allow light to pass there throughfrom below sensor 100, reflect off reflector 137, and exit again throughtransmission window 119. It will be noted, that in some embodiments,base layer 110 is entirely formed from a transparent material.

The thickness of first spacer layer 120 along with the adhesive thatbonds it to the adjacent layers defines a first depth 153 between thebase layer 110 and the reflector 137. In some embodiments, first depth153 is approximately 0.004 inches deep. First depth 153 is used todetermine the effective path length for optical hemoglobin measurement,discussed in greater detail below. The combined thicknesses of the firstspacer 120, second spacer 130, and adhesive layers that bind themtogether define a second depth 155.

FIG. 6 depicts a longitudinal cross-sectional view of an embodiment ofsensor 100 that includes a deposit of porous, reflective material 161 onbase layer 110. In some embodiments, a deposit of porous, reflectivematerial 161 may be screen-printed directly on top of transmissionwindow 119. The deposit of porous, reflective material 161 may bedisposed between at least two of electrodes 115-117. In embodimentsincluding a deposit of porous, reflective material 161, only a firstspacer layer 120 need be used. Accordingly, these embodiments may omitsecond spacer layer 130 including bridge 136 and modify the thickness ofthe first spacer layer to be approximately 0.013 inches. The deposit ofporous, reflective material 161 should be made with a material that canabsorb the liquid sample and whose reflectance changes as the sample isabsorbed. In some embodiments, the deposit of porous, reflectivematerial 161 is formed from a porous paper, porous ink, or polymerfilter material. The use of this embodiment in making an opticalhemoglobin measurement will be discussed in greater detail below.

FIG. 7 depicts an embodiment of an analyzer configured to receive andanalyze a sample on sensor 100. Sensor 100 is configured in size andshape to be insertable into an analyzer 200. The analyzer 200 mayinclude a housing 205 configured in size and shape to be used on atabletop or lab bench. In some embodiments, the housing 205 may beconfigured for hand held use. Housing 205 includes a display 207 thatdisplays instructions and sample results to an operator. In someembodiments, the display 207 is an interactive display, such as a touchscreen, which enables an operator to view, set, or select variousanalysis parameters and view sample results. In some embodiments, theanalyzer 200 comprises an input device, such as a keyboard, soft or hardbuttons, a mouse, or any other suitable input device which allows anoperator to interact with the analyzer 200.

Housing 205 includes a sensor port 208 through which a sensor supportstructure 250 extends. Sensor port 208 may further be configured in sizeand shape to receive the first end 101 of sensor 100 through housing205. The analyzer 200 may be configured with a single sensor port 208 toaccept and analyze a single sensor 100 or with a plurality of sensorports 208 to accept a plurality of sensors 100. A suitable analyzer foruse in sampling blood lead levels is described in U.S. Pat. No.5,873,990, entitled “Handheld Electromonitor Device,” and in U.S. patentapplication Ser. No. 13/790,154, the entire contents of which are hereinincorporated by reference.

As shown in FIG. 7, sensor support structure 250 extends through housing205 at sensor port 208. Sensor support structure 250 includes a supportsurface 251 on which sensor 100 rests when inserted into analyzer 200.Further, sensor support structure 250 may further comprise sensor guides253 a, 253 b, each of which may be configured to extend upward fromsupport surface 251 and form a wall oriented in a direction parallel toa longitudinal axis of sensor 100 when sensor 100 is inserted intoanalyzer 200. Sensor guides 253 a, 253 b may further include anoverhanging portion that covers at least a portion of a top surface ofsensor 100 when sensor 100 is inserted. Sensor guides 253 a, 253 b andsupport surface 251 thus provide correct orientation and stability forsensor 100 as it is inserted into analyzer 200.

FIG. 8 depicts an embodiment of sensor support structure 250 removedfrom housing 205 for ease of description. Sensor support structure 250includes an external end 258 (the portion extending through the housing205 in FIG. 4) and an internal end 259, which is contained within thehousing 205. Support surface 251 may comprise a substantially flatsurface sized and shaped to support sensor 100 at an orientation that issubstantially parallel to a surface on which analyzer 200 is resting.Sensor guides 253 a, 253 b extend upward from lateral sides of supportsurface 251. Internal end 259 may include a plurality of sensor contacts261 disposed within an electrical contact structure 260. The pluralityof sensor contacts 261 are positioned to contact the contacts 111-114 ofsensor 100 when sensor 100 is inserted into sensor port 208. In someembodiments there is at least one sensor contact 261 for each ofcontacts 111-114 of sensor 100. In some embodiments, more than onesensor contact 261 may contact one of contacts 111-114. For example, insome embodiments, two sensor contacts 261 each make an electricalconnection with contact 114 of sensor 100. In this way, contact 114completes a circuit which signals analyzer 200 that a sensor 100 hasbeen inserted.

Sensor support structure 250 may also comprise an aperture 255 which isformed as a hole extending through support surface 251. In someembodiments, aperture 255 may be filled with a window 257. Aperture 255is positioned on the sensor support structure to correspond to thetransmission window 119 formed in the base layer 110 of the sensor 100.In this way, when a sensor 100 is inserted into the sensor port 208, anoptical path is created between the transmission window 119 and theaperture 255 through which an optical signal can pass.

The window 257 is made from a scratch resistant material that permitslight of the wavelengths discussed below to pass there through. In someembodiments, window 257 may comprise glass, transparent polycarbonateplastic, or other suitable material. Some embodiments may advantageouslyuse a material with a high index of refraction, for example, materialswith a refractive index greater than 1.4. When materials with higherindices of refraction are used, incoming light that enters the window ata shallow angle will be refracted at a steeper angle, thus contactingand reflecting off reflector 137 at the steeper angle. The anglesdiscussed in this paragraph are measured between the ray of light and anaxis normal to the surface of reflector 137. Angles approaching 0degrees are considered steeper while angles approaching 90 degrees areconsidered shallower. In some embodiments, the window 257 may comprise asapphire window.

An optical system 300 is also shown in FIG. 8 and is disposedsubstantially below sensor support structure 250 and within housing 205.In some embodiments, optical system 300 comprises an optical systemhousing 303. Aperture 255 extends through support surface 251 and intooptical system housing 303.

Optical system 300 is now described in greater detail with reference toFIGS. 9-10. FIG. 9 depicts a cross-sectional view of an embodiment of anoptical housing 303 and the various components that may be containedtherein. Optical system 300 includes first and second light sources 321,322, collection lens 313, and detector 311 all disposed within housing303. In some embodiments, the optical system 300 may include only asingle light source (for example, light source 321 or light source 322),or more than two light sources. However, the following descriptionpresents a non-limiting example that includes two light sources. Firstand second light sources 321, 322 may also include correspondingelectrical connections 321 a, 322 a for powering and controlling thefirst and second light sources. In some embodiments, the optical system300 may include only a single light source. In some embodiments, thefirst and second light sources 321 and 322 comprise a single LED or LEDchip. In some embodiments, the first and second light sources 321 and322 comprise one or more LED chips. In some embodiments, the first andsecond light sources 321 and 322 comprise 4 LED chips locatedsymmetrically about the longitudinal axis of the first channel 315.

A first source channel 315 may be formed as a hole extending throughoptical system housing 303. The first source channel 315 may extendbetween the first light source 321 and the aperture 255. In someembodiments, first source channel 315 includes a narrow portion 315 a,wherein the narrow portion 315 a comprises a diameter smaller than thediameter of first source channel 315 where the first light source 321 isdisposed. First light source 321 is disposed within first channel 315and oriented so that a central axis of the light emitted from firstlight source 315 is substantially coaxial with a longitudinal axis offirst channel 315. In some embodiments, the central axis of the lightemitted from the first light source 315 is not coaxial with thelongitudinal axis of the first channel, and is arranged so that at leasta portion of the emitted light travels the length of the longitudinalaxis of the first channel 315 and exits through the aperture 255. Insome embodiments, first light source 321 is disposed within first sourcechannel 315 at a position below narrow portion 315 a. A second sourcechannel 317 may be formed as a hole extending through optical systemhousing 303 similar to the first source channel 315. The second sourcechannel 317 may extend between the second light source 322 and aperture255. In some embodiments second source channel 317 includes a narrowportion 317 a, wherein the narrow portion 317 a comprises a diametersmaller than the diameter of second source channel 317 at the locationof the second light source 322. Second light source 322 is disposedwithin second channel 317 and oriented so that a central axis of thelight emitted from second light source 317 is coaxial with alongitudinal axis of second channel 317. In some embodiments, secondlight source 322 is disposed within second source channel 317 at aposition below narrow portion 317 a. In some embodiments, the firstchannel 315 and the second channel 317 may be oriented such that thelongitudinal axes of the first channel 315 and the second channel 317are perpendicular to each other. In some embodiments, the longitudinalaxes of the first channel 315 and the second channel 317 may intersect,forming an acute or obtuse angle.

A collection channel 318 is also disposed within housing 303 and isformed as a hole extending between aperture 255 and a bottom surface ofhousing 303. Collection channel 318 is disposed below aperture 255 andhas a longitudinal axis that extends in a direction normal to the planeof aperture 255. Detector 311 is disposed in or below collection channel318 on the end of collection channel 318 opposite aperture 255. Detector311 may comprise a photo diode with an integral amplifier, aphotomultiplier or another optical detector capable of measuring lightintensity. In some embodiments, a collection lens 313 is disposed incollection channel 318 between detector 311 and aperture 255. Collectionchannel 318 or optical housing 303 may include a mounting structure forsecuring collection lens 313. Collection lens 313 is oriented andconfigured in size and shape to focus light traveling from the aperture255, through collection channel 318 onto detector 311. An angle α isformed between each of the longitudinal axes of first and second sourcechannels 315, 317 and the longitudinal axis of collection channel 318.In other words, α is the angle between how a light source 321, 322 isaimed and an axis extending normal to the detector 311. In someembodiments, α is approximately 45°. In some embodiments, a isapproximately 10°, 15°, 20°, 25°, 30°, 35°, 40°, 50°, 55°, 60°, 65°, 70°75°, 80°, 85°, 90°, or more, or any angle there between. It should beunderstood, however, that the value of α affects the reflectance oflight emitted by the first light source 321 and the second light source322 as the light passes through the aperture 255 and through the samplein the first sample reservoir space 123. In some embodiments, the angleα of the longitudinal axes of the first channel 315 and the secondchannel 317 may be the same as each other, or may be different. Forexample, the angle α for the first source channel 315 may beapproximately 45°, and the angle α for the second source channel 317 maybe other than 45°. It will be noted that while first and second channels315, 317 and collection channel 318 have all been depicted as lying inthe same plane in FIG. 9, this may not be the case for all embodiments.

In some embodiments, a washer 254 may be an aperture, such as astructure including a center hole 254a may be disposed below or attachedto the underside of window 257 in aperture 255. Washer 254 with centerhole 254a may be configured to narrow the beam of light passing throughaperture 255. In some embodiments, washer 254 is made from plastic,rubber, or metal and may be finished with a flat (non-glossy)non-reflective surface.

Some of the components of an embodiment of an optical system 300 can beseen more clearly in FIG. 10, which depicts a perspective view of anembodiment of an optical system 300 with the optical system housing 303removed. Optical system 300 includes first light source 321 and secondlight source 322. In some embodiments, the first and second lightsources 321, 322 include integrated lenses 325, 326 which are configuredto focus the light emitted through aperture 255 and onto reflector 137of sensor 100. Each of the first and second light sources 321, 322 maycomprise a plurality of LEDs positioned on a printed circuit board. Insome embodiments, each of the first and second light sources 321, 322comprise four LEDs positioned on a printed circuit board. It will,however, be understood by one of skill in the art that a single LED orother light source may be used. Additionally, in some embodiments asingle light source 321 may be used or more than two light sources maybe used.

The light sources 321, 322 may be configured to emit light with anapproximately 405 nm wavelength, the benefits of which will be discussedbelow. In some embodiments, the wavelength may be about 410 nm. In someembodiments, the wavelength may be from about 350 nm to about 450 nm. Insome embodiments, the wavelength can be between 250 nm and 950 nm. Itwill be understood by one of skill in the art that other wavelengths oflight can be used.

In one embodiment, optical system 300 includes the electronic componentsillustrated schematically in FIG. 11. A time base generator or clock 351is electrically connected to a current source 353 such that an output ofclock 351 is an input to current source 353. Current source 353 is thenelectrically connected to first and second light sources 321, 322.Detector 311 is electrically connected to an amplifier 355 such that anoutput of the detector 311 is an input of the amplifier 355. Amplifier355 is further electrically connected to a demodulator 357 such that anoutput of amplifier 355 is a first input of demodulator 357. Demodulator357 is also electrically connected to the clock 351 such that an outputsignal from clock 351 is a second input of demodulator 357. In someembodiments, demodulator 357 is electrically connected to a low passfilter 359 such that an output of demodulator 357 is an input of lowpass filter 359. Low pass filter 359 may then be electrically connected,either directly or indirectly, to a processor 201. Processor 201 may beconnected to and control the clock 351. In some embodiments, more thanone (for example, two), low pass filters may be used. For example, afirst low pass filter may have a larger time constant (in other words, aslower response) and be used during measurement of a filled sensor, anda second low pass filer may have a shorter time constant (in otherwords, a faster response) and be used during a reference measurementtaken while the sensor is being inserted (as will be described below ingreater detail in reference to FIG. 16). The processor 201 may selectbetween the two low pass filters in this example. Processor 201 maycontrol all the components depicted in FIG. 11, and may further controlthe operations of the analyzer 200. Processor 201 may comprise more thanone processor.

The arrangement of components shown in FIG. 11 and described above mayprovide improved lock-in signal processing in some embodiments ofoptical system 300. The output signal of clock 351 may be used to drivecurrent source 353 at a particular frequency. Current source 353 willthen, accordingly, drive the first and second light sources 321, 322such that they flash at the frequency indicated by clock 351. The lightfrom first and second light sources 321, 322 passes through the sampleand is reflected off reflector 137. At least a portion of the lightreflected off reflector 137 travels through the sample again, toward theaperture 255, is received by detector 311. The detector 311 converts theoptical signal into an electrical output signal. The output of detector311 is amplified at amplifier 355 and fed as a first input todemodulator 357. Demodulator 357 also receives, as a second input, theoutput of the clock. Accordingly, demodulator 357 is able to distinguishthe portion of light received at the detector 311 due to light emittedfrom light sources 321, 322 at the frequency of the clock 351, or at afrequency having a known deviation from the frequency of the clock 351from light received at the detector 311 from other ambient sources,which has a frequency other than that of the clock 351 or other than theknown deviation from the frequency 351 of the clock. The demodulatorremoves substantially any signal from the electrical output which doesnot correspond to light emitted at the frequency of the clock 315. Insome embodiments, the LEDs are flashed at approximately 100 Hz, 500 Hz,1 kHz, 1.5 kHz, 2 kHz, 5 kHz, 10 kHz, 50 kHz, or more, or any valuethere between; it will be understood by one of skill in the art andaccording to the principles taught here, that other frequencies may beused without departing from the scope of this disclosure.

Sensor 100 and analyzer 200 can be used to make simultaneous measurementof blood lead and hemoglobin concentration as follows. First, a sensor100 is inserted into analyzer 200 at port sensor port 208. Contact 114comes into contact with sensor contacts 261 of analyzer 200 completing acircuit within analyzer 200 that signals that sensor 100 has beeninserted. Analyzer 200 may determine whether a sensor has been insertedaccording to the methods disclosed in U.S. patent application Ser. No.13/790,154, entitled “Apparatus and Method for Analyzing MultipleSamples,” which has been previously incorporated by reference above.

Analyzer 200 may further perform routines to ensure that the sensor 100that has been inserted has not previously been used. Accordingly,analyzer 200 may check to ensure that the sensor 100 has been wetted. Ifa wetted sensor has been inserted, analyzer 200 may provide an errormessage indicating that a previously used sensor has been inserted. Thiswill prompt the user to discard the old sensor and insert a fresh one.This determination may also be made with the methods disclosed in U.S.patent application Ser. No. 13/790,154. As used herein, the term“wetting the sensor” is used to indicate introducing a sample, such as ablood sample that may be prepared with a reagent, to the sensor and a“wetted sensor” indicates a sensor wherein the sample has beenintroduced.

At this point, analyzer 200 may provide a user with prompts on display207 giving the user an option of which tests should be performed. Theuser may select blood hemoglobin concentration, blood leadconcentration, or both. In another embodiment, the sensor may beprogramed to automatically test for both blood hemoglobin and blood leadconcentration and no prompts will provided to the user.

If a blood hemoglobin concentration will be performed, analyzer 200 willtake an optical reference measurement, for example, of the empty sensor100 prior to introduction of the prepared sample. This referencemeasurement will be discussed in greater detail below.

Analyzer 200 may then prompt the user to introduce the prepared sampleinto the sensor 100 that has been inserted into analyzer 200. The usermay prepare the sample by mixing the blood sample with a solution ofhydrochloric acid, which reagent prepares the sample for anelectrochemical lead concentration measurement as discussed elsewhere.The user may then transfer the prepared sample to sensor 100 with apipette or dropper, introducing the prepared sample at sample inlet 141filling sample reservoir 151. Analyzer 200 may again check whether thesensor 100 has been wetted using the methods indicated above. Onceanalyzer 200 determines that the sample has been introduced, hemoglobinconcentration and blood concentration analysis may begin.

Blood hemoglobin concentration analysis will be described first;however, this analysis may proceed simultaneously with the blood leadconcentration analysis described below.

The optical absorbance of blood treated with hydrochloric acid dependson the hemoglobin concentration of the sample. For example, FIG. 12depicts three correlation curves obtained experimentally using acommercial UV/Vis spectrophotometer. As shown, absorbance measurementsof prepared samples were taken at three light wavelengths, 410 nm, 520nm, and 700 nm, each yielding a substantially linear relationshipbetween the hemoglobin concentration (measured in g/dL) and theabsorbance of the sample. This linear relationship can be describedusing the Beer-Lambert Law:

Concentration=Absorbance/(ε×path)   [1]

Absorptivity, ε, is a property of hemoglobin. Path is the length of thesample through which a beam of light is passed and can be obtained fromthe linear dimension of the cuvette in which the sample is contained,and the Absorbance can be calculated as follows:

Absorbance=−log(I/I ₀)   [2]

I is the measured intensity of light passing through the sample cuvetteand I₀ is the intensity of a reference beam, which can be obtained bypassing the light through a reference cuvette. The reference cuvette maybe empty or may contain a liquid that does not include any hemoglobin,for example.

These general principles may be modified and implemented in analyzer 200as follows to allow analyzer 200 to optically determine the hemoglobinconcentration of the prepared sample using a reflectance measurement.Optical system 300 may be configured with the various componentsdiscussed above to allow it to measure the intensity of the lightreflected off reflector 137 or off a deposit of porous, reflectivematerial 161. The discussion below will provide an example of a sensor100 including a reflector 137; similar principles apply by analogy, toembodiments of sensor 100 including a deposit of porous, reflectivematerial 161. As shown in FIG. 13, first and second light sources 321,322 emit light upward toward the reflector 137 of sensor 100. In someembodiments, first and second light sources 321 and 322 emit lightsimultaneously, and in some embodiments, the first and second lightsources 321 and 322 alternately emit light. In some embodiments, firstand second light sources 321, 322 are pulsed at approximately 1 kHz asdescribed above. To reach reflector 137, the light travels upwardthrough the aperture 255 in optical system housing 303 and window 257.The light continues through the transmission window 119 of base layer110 of sensor 100 and passes through the sample in sample reservoir 151until some fraction of it is diffusely reflected downwards off reflector137. A portion of the reflected light travels out through the aperture255 into the collection channel 318, where it is focused with collectionlens 313 towards detector 311. Detector 311 measures the intensity ofthe light received. The intensity signal may then be converted to anelectrical signal and input to processor 201 of analyzer 200 and used tocalculate the hemoglobin concentration.

First depth 153, defined by the thickness of first spacer layer 120 anddiscussed above with reference to FIGS. 5A and 5B, should besufficiently thin so as to ensure that some light is reflected back outof sensor 100. If first depth 153 is overly deep, substantially all ofthe light entering sensor 100 through transmission window 119 will beabsorbed by the sample and nothing will be reflected and measured. Thiseffect can be minimized by ensuring that first depth 153 is sufficientlythin, for example, about 0.004″ or by increasing the intensity of thelight emitted from first and second light sources 321, 322.

As shown in FIG. 14, optics system 300 may first take a reference scanof sensor 100 before the sample has been introduced to generatereference intensity measurement, I₀. The sample may then be introducedand optical system 300 can take a second measurement, yielding themeasured intensity of light reflected through the sample, I. Reflectancecan then be calculated using the following equation:

Reflectance=−log(I/I ₀)   [3]

The same sensor 100 can be used for each measurement, with a firstmeasurement being taken while the sensor is empty and a secondmeasurement taken after the sample is introduced to the sensor. In otherembodiments, however, two sensors, with similar dimensions and opticalcharacteristics, may be used: a first empty sensor and a second filledsensor; this will, however, yield less accurate results due tovariations in sensor dimensions due to manufacturing and variations ofsensor positioning within the analyzer.

In some embodiments, analyzer 200 with optical system 300 may further becalibrated to account for internally reflected stray light or anycontribution from any fluorescence produced by the substrate of sensor100 to achieve more precise and consistent results. As shown in FIG. 15,some light reflected off internal components of optical system 300 maybe received at detector 311. To correct for this stray light,reflectance measurement may be adjusted as follows:

Reflectance=−log((I−I _(stray))/(I₀ −I _(stray)))   [4]

The internally reflected stray light, I_(stray), can be estimated bytaking a measurement with no sensor in place. This measurement can thenbe subtracted from the measurements taken of the filled and unfilledsensor.

Alternatively, the internally reflected stray light, I_(stray), can beestimated by measuring the reflected signal from a light-absorbing blacksurface on sensor 100 as sensor 100 is inserted into or withdrawn fromanalyzer 200. This embodiment is depicted in FIG. 16, which shows howthe measured intensity of reflected light varies as sensor 100 isinserted into analyzer 200. As the sensor is inserted, it moves acrossthe light beam emitted from optical system 300. As shown in FIG. 16, asthe beam passes over a light-absorbing black surface 118 the measuredintensity falls to a level representing the internally reflected straylight, I_(stray). Once the sensor is fully inserted, the beam is focusedon reflector 137 yielding a value representing I if the sensor is filledor I₀ if the sensor is empty. In some embodiments, the light-absorbingblack surface 118 may be the carbon of the counter electrode 117. Inother embodiments it may be a coating applied to the bottom surface ofbase layer 110. The width of the light-absorbing black surface 118 maybe adjusted to provide for a more accurate measurement of I_(stray) asthe sensor is inserted. In some embodiments, the width of thelight-absorbing black surface 118 is between 1-5 millimeters. In someembodiments, the width of the light-absorbing black surface 118 isbetween 3.5-4 mm. In another embodiment, the internally reflected straylight, I_(stray), is estimated by taking the difference between the twomethods previously described—in other words, by taking the differencebetween a measurement of a reference cuvette and a measurement of alight-absorbing black surface. Experimentally, this difference has beenfound to provide an accurate estimate of the internally reflected straylight, I_(stray).

In some embodiments, measurements for I₀ and I_(stray) can be obtainedby mechanically moving a sample white surface and a sample black surfaceinto contact with the optical system and measuring the reflectance.

The reflectance measured and calculated using equation [3] or correctedequation [4] above does not give a linear relationship betweenreflectance and hemoglobin concentration of the sample. Nonetheless, anonlinear calibration curve can be calculated that will allow analyzer200 to determine hemoglobin concentration from reflectance using themeasurement of reflectance described above. Two non-linear examplecalibration curves are shown in FIG. 17. In the figure, calibrationcurves for light with wavelengths of 405 nm and 625 nm are shown assecond degree polynomials. One of skill in the art will appreciate thatother functions, other than second degree polynomials, may be used.

The wavelength of light in the measurement may also affect the accuracyof the analysis. As noted above, first and second light sources 321, 322may use light with a wavelength between 250 nm and 950 nm. The inventorshave observed that the absorbance or reflectance of a blood sampletreated with hydrochloric acid change over time. For example, a sampletreated with hydrochloric acid and measured immediately may yield adifferent absorbance or reflectance value than the same sample measuredagain 10 minutes later. It has further been observed that change overtime of the absorbance or reflectance of a sample is also affected bythe wavelength of light used to make the measurement. These results canbe seen in FIG. 18 which shows the measured absorbance of a treatedsample measured with wavelengths of light between 350 nm and 450 nm withmeasurements taken every minute for ten minutes. As can be seen in FIG.18, at wavelengths less than approximately 405 nm, the measuredabsorbance decreases over time. At wavelengths greater thanapproximately 405 nm, this trend reverses and the measured absorbanceincreases over time. Importantly, the inventors have observed that atapproximately 405 nm, the absorbance does not change over time,indicating an isosbestic point. Accordingly, because the time betweenpreparation of the sample and measurement of the sample may vary, it ispreferred to configure analyzer 200 to measure with light ofapproximately 405 nm. In some embodiments, analyzer 200 may measure at anumber of wavelengths in addition to 405 nm and use the additionalinformation to improve the precision of the measurement.

The apparatus and methods disclosed herein for making an opticalhemoglobin measurement of a treated blood sample may be modified toallow for measurement with different geometries. For example, throughoutthis application, reference has been made to optically measuring forhemoglobin concentration using a light source and detector positionedgenerally below the sensor wherein the light from the light sourcepasses upward through the sample and is reflected back down to thedetector. This is merely exemplary. One of skill in the art willunderstand, according to the principles herein disclosed, that the lightsource and detector could be positioned generally above the sensor. Insome embodiments, the light source may be positioned on one side of thesensor and the detector could be positioned on the opposite side of thesensor such that the light emitted travels through a transparent portionof the lid or through a hole in the lid, through the sample, and througha transparent portion on the base of the sensor.

In addition to the blood hemoglobin concentration previously described,analyzer 200 may also be configured to simultaneously measure blood leadconcentration using the same sensor 100. Blood lead concentrationanalysis can be performed electrochemically using sensor 100 andanalyzer 200 as described in U.S. Pat. No. 5,368,707, entitled“Convenient Determination of Trace Lead in Whole Blood and OtherFluids,” the entire contents of which is herein incorporated byreference, and U.S. Pat. No. 5,468,366, entitled “Colloidal-GoldElectrosensor Measuring Device,” mentioned previously above.

Upon completion of the hemoglobin and lead concentration analyses,analyzer 200 may display the results of the analysis to the user viadisplay 207. Alternatively, results may be stored, sent to an externalcomputer, or printed.

Accordingly, the embodiments and principles described above may be usedto measure the lead and hemoglobin concentrations in a blood samplesimultaneously, using a single sensor and analyzer.

Example Hemoglobin Measurement

A sensor and analyzer incorporating the above-described principles foroptically measuring hemoglobin has been developed and tested yieldingthe following results. The analyzer was configured to calculatehemoglobin concentration using the 405 nm calibration curve shown inFIG. 15:

y=−0.0028x ²+0.1489x+0.1081   [5]

where y represents the reflectance calculated as−log((I−I_(stray))/(I₀−I_(stray))), and x represents the hemoglobinconcentration with units of g/dL.

Forty whole blood samples were obtained by venipuncture and stored atrefrigerated temperature for less than 72 hours prior to analysis. Fiftymicroliters of blood sample were added to one tube of MagellanDiagnostics LeadCare treatment reagent, mixed thoroughly for one minuteand introduced into a sensor. The light intensity at 405 nm reflectedfrom the sensor was measured before (I₀) and after (I) the sample wasintroduced. The concentration of hemoglobin was determined using thecalibration curve presented above. The same samples were tested using anInstrumentation Laboratories GEM Premier 4000 co-oximeter to obtain areference value for comparison. As shown in FIG. 18, there is anexcellent correlation between the hemoglobin concentration determinedusing the principles herein disclosed and the reference value.

The foregoing description details certain embodiments of the systems,devices, and methods disclosed herein. It will be appreciated, however,that no matter how detailed the foregoing appears in text, the systems,devices, and methods can be practiced in many ways. As is also statedabove, it should be noted that the use of particular terminology whendescribing certain features or aspects of the embodiments disclosedherein should not be taken to imply that the terminology is beingre-defined herein to be restricted to including any specificcharacteristics of the features or aspects of the technology with whichthat terminology is associated.

It will be appreciated by those skilled in the art that variousmodifications and changes may be made without departing from the scopeof the described technology. Such modifications and changes are intendedto fall within the scope of the embodiments. It will also be appreciatedby those of skill in the art that parts included in one embodiment areinterchangeable with other embodiments; one or more parts from adepicted embodiment can be included with other depicted embodiments inany combination. For example, any of the various components describedherein and/or depicted in the Figures may be combined, interchanged orexcluded from other embodiments.

With respect to the use of substantially any plural and/or singularterms herein, those having skill in the art can translate from theplural to the singular and/or from the singular to the plural as isappropriate to the context and/or application. The varioussingular/plural permutations may be expressly set forth herein for sakeof clarity.

It will be understood by those within the art that, in general, termsused herein are generally intended as “open” terms (e.g., the term“including” should be interpreted as “including but not limited to,” theterm “having” should be interpreted as “having at least,” the term“includes” should be interpreted as “includes but is not limited to,”etc.). It will be further understood by those within the art that if aspecific number of an introduced claim recitation is intended, such anintent will be explicitly recited in the claim, and in the absence ofsuch recitation no such intent is present. For example, as an aid tounderstanding, the following appended claims may contain usage of theintroductory phrases “at least one” and “one or more” to introduce claimrecitations. However, the use of such phrases should not be construed toimply that the introduction of a claim recitation by the indefinitearticles “a” or “an” limits any particular claim containing suchintroduced claim recitation to embodiments containing only one suchrecitation, even when the same claim includes the introductory phrases“one or more” or “at least one” and indefinite articles such as “a” or“an” (e.g., “a” and/or “an” should typically be interpreted to mean “atleast one” or “one or more”); the same holds true for the use ofdefinite articles used to introduce claim recitations. In addition, evenif a specific number of an introduced claim recitation is explicitlyrecited, those skilled in the art will recognize that such recitationshould typically be interpreted to mean at least the recited number(e.g., the bare recitation of “two recitations,” without othermodifiers, typically means at least two recitations, or two or morerecitations). Furthermore, in those instances where a conventionanalogous to “at least one of A, B, and C, etc.” is used, in generalsuch a construction is intended in the sense one having skill in the artwould understand the convention (e.g., “a system having at least one ofA, B, and C” would include but not be limited to systems that have Aalone, B alone, C alone, A and B together, A and C together, B and Ctogether, and/or A, B, and C together, etc.). In those instances where aconvention analogous to “at least one of A, B, or C, etc.” is used, ingeneral such a construction is intended in the sense one having skill inthe art would understand the convention (e.g., “a system having at leastone of A, B, or C” would include but not be limited to systems that haveA alone, B alone, C alone, A and B together, A and C together, B and Ctogether, and/or A, B, and C together, etc.). It will be furtherunderstood by those within the art that virtually any disjunctive wordand/or phrase presenting two or more alternative terms, whether in thedescription, claims, or drawings, should be understood to contemplatethe possibilities of including one of the terms, either of the terms, orboth terms. For example, the phrase “A or B” will be understood toinclude the possibilities of “A” or “B” or “A and B.”

While various aspects and embodiments have been disclosed herein, otheraspects and embodiments will be apparent to those skilled in the art.The various aspects and embodiments disclosed herein are for purposes ofillustration and are not intended to be limiting.

What is claimed is:
 1. A method for measuring an analyte in a liquidsample, the method comprising: flowing a liquid sample into a reservoirof a sensor, the reservoir comprising a first depth and a second depththat is greater than the first depth; illuminating the liquid sample inthe reservoir through a transparent portion of the sensor; measuring areflectance of the liquid sample; and computing a measurement of theanalyte using the measured reflectance.
 2. The method of claim 1,wherein the reflectance is measured by measuring light reflected from areflective surface in the sensor.
 3. The method of claim 2, wherein thereflective surface is positioned within the reservoir.
 4. The method ofclaim 3, wherein the reflective surface is positioned within the firstdepth of the reservoir.
 5. The method of claim 1, wherein thereflectance is computed by comparing a measured intensity to a referenceintensity.
 6. The method of claim 5, further comprising measuring thereference intensity by: illuminating an empty sensor; and measuring areflectance of the empty sensor.
 7. The method of claim 5, furthercomprising: measuring internally reflected stray light by detecting theintensity of light reflected from a light absorbing surface on thesensor; subtracting the measured internally reflected stray light fromthe reference intensity and the measured intensity.
 8. The method ofclaim 7, wherein measuring the internally reflected stray lightcomprises measuring the internally reflected stray light by detectingthe intensity of light reflected from a light absorbing surface on thesensor as the sensor is inserted into or withdrawn from an analyzer. 9.The method of claim 1, further comprising measuring a second analyte inthe liquid sample using an electrode positioned within the reservoir ofthe sensor.
 10. The method of claim 9, wherein the electrode ispositioned within the second depth of the reservoir.
 11. A sensor formeasuring an analyte comprising: a reservoir having a top surface and abottom surface, at least a portion of the top surface comprising areflector and at least a portion of the bottom surface comprising atransparent portion; wherein the reservoir comprises a first portionhaving first depth between the bottom surface of the reservoir and afirst portion of the top surface of the reservoir, and a second portionhaving a second depth between the bottom surface of the reservoir and asecond portion of the top surface of the reservoir, wherein the firstdepth is different from the second depth.
 12. The sensor of claim 11,further comprising: at least one electrode disposed on the bottomsurface of the reservoir; and at least one electrical contact inelectrical communication with the at least one electrode.
 13. The sensorof claim 12, wherein the at least one electrode is disposed in thesecond portion of the reservoir.
 14. The sensor of claim 12, wherein afirst of the at least one electrodes comprises a colloidal gold deposit.15. The sensor of claim 14, wherein a second of the at least oneelectrode comprises a carbon electrode disposed on the base layer. 16.The sensor of claim 11, further comprising a sample inlet and a vent,each formed as a hole extending through the lid.
 17. The sensor of claim11, wherein a thickness of the sensor is less than about 0.05 inches.18. The sensor of claim 11, wherein the transparent portion comprises apolycarbonate material.